Monte Carlo Analysis of Neck Linker Extension in Kinesin Molecular Motors

Abstract

Kinesin stepping is thought to involve both concerted conformational changes and diffusive movement, but the relative roles played by these two processes are not clear. The neck linker docking model is widely accepted in the field, but the remainder of the step – diffusion of the tethered head to the next binding site – is often assumed to occur rapidly with little mechanical resistance. Here, we investigate the effect of tethering by the neck linker on the diffusive movement of the kinesin head, and focus on the predicted behavior of motors with naturally or artificially extended neck linker domains. The kinesin chemomechanical cycle was modeled using a discrete-state Markov chain to describe chemical transitions. Brownian dynamics were used to model the tethered diffusion of the free head, incorporating resistive forces from the neck linker and a position-dependent microtubule binding rate. The Brownian dynamics and chemomechanical cycle were coupled to model processive runs consisting of many 8 nm steps. Three mechanical models of the neck linker were investigated: Constant Stiffness (a simple spring), Increasing Stiffness (analogous to a Worm-Like Chain), and Reflecting (negligible stiffness up to a limiting contour length). Motor velocities and run lengths from simulated paths were compared to experimental results from Kinesin-1 and a mutant containing an extended neck linker domain. When tethered by an increasingly stiff spring, the head is predicted to spend an unrealistically short amount of time within the binding zone, and extending the neck is predicted to increase both the velocity and processivity, contrary to experiments. These results suggest that the Worm-Like Chain is not an adequate model for the flexible neck linker domain. The model can be reconciled with experimental data if the neck linker is either much more compliant or much stiffer than generally assumed, or if weak kinesin-microtubule interactions stabilize the diffusing head near its binding site.

Neck-Linker Length Dependence of Processive Kinesin-5 Motility

“…19,25,26 For a set of particular Kinesin-1 and Kinesin-2 constructs, it was found that processivity decreases with increasing neck-linker length, while shortening of the native neck linker by one amino acid for that motor abolishes processivity. 24,[27][28][29] A continuation of this work showed similar effects for Kinesin-1/ Kinesin-2/Kinesin-3 and Kinesin-7 constructs. 25 In another study, 26 the insertion of up to 26 prolines in the neck linker of human Kinesin-1 was reported to not only slightly decrease run length under lowionic-strength conditions but also strongly decrease motor speed and stall force.…”

“…22,23 When modeled as a simple flexible polymer, the tension in the neck linker at a given end-to-end distance and, therefore, the strength of the head-head communication would scale inversely with contour length, that is, number of amino acids. 24 Data from three recent studies support such a simple general model of head-head communication. 19,25,26 For a set of particular Kinesin-1 and Kinesin-2 constructs, it was found that processivity decreases with increasing neck-linker length, while shortening of the native neck linker by one amino acid for that motor abolishes processivity.…”

Interhead tension determines processivity across diverse N-terminal kinesins

“…The role of the neck linker domain in tethered diffusion of the free head was recently investigated by our group using Brownian dynamics simulations (39). If the neck linker is modeled as a worm-like chain, the neck linker is expected to strongly hinder the diffusion of the tethered head to the next binding site, and a large force between the heads (>30 pN) is expected in the two-head bound state.…”

Stepping molecular motor amid Lévy white noise

“…The elastic properties of a linker structure in typical molecular motors, like kinesins, have been formerly addressed in experimental studies and molecular dynamics simulations using g rom acs software [68,69], Based on accumulated evidence, it is unlikely that a molecular machine would respond instantaneously to an elongation with an elastic counteraction that resets the linker structure to its equilibrium conformation of length a or shorter (see Fig. 2).…”